Herbicide degradation and copper complexation by bacterial mixed cultures from a vineyard stormwater basin
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The use of stormwater basins as constructed wetlands for the bioremediation of agricultural runoff waters contaminated with pesticides has great potential. The structure and dynamics of the bacterial community in such system, and its function with respect to contaminant removal, remain to be investigated in detail.
Materials and methods
The bacterial component of sediment collected from a vineyard stormwater basin (in Rouffach, France) that pooled incoming runoff water containing pesticide and copper, was investigated by enumeration of cultivable bacteria on sediment extract solid medium and by liquid enrichment cultures from sediment, in the presence of glyphosate, diuron, 3,4-dichloroaniline and copper. Its structure, as a function of sediment location, depth, rhizospheric status and the presence of contaminants, was studied by temporal temperature gradient electrophoresis. Cultures obtained by enrichment were screened by RISA and RFLP and the ability of different cultures for contaminant mitigation was evaluated by the chrome azurol S method (copper complexation) and HPLC (glyphosate, diuron and 3,4-dichloroaniline degradation). The composition of the mixed cultures with the highest potential with regard to degradation of glyphosate, diuron and 3,4-dichloroaniline and copper complexation were evaluated by sequence analysis of cloned PCR-amplified 16S rRNA gene fragments obtained from enrichment cultures.
Results and discussion
The bacterial community structure of sediment showed differences depending on sampling location, sediment depth and sampling date. Spiking with a cocktail of concentrated glyphosate, diuron, 3,4-dichloroaniline and copper altered the bacterial community structure, but rhizospheric samples were less affected. RISA and RFLP analysis differentiated 98 distinct cultures, 28 of which were able to complex copper, and three, 35 and seven were able to degrade glyphosate, diuron and 3,4-dichloroaniline, respectively. Sequencing of cloned 16S rRNA gene fragments amplified from faster-growing rhizospheric mixed culture 106, selected as the most efficient in complexing copper and degrading glyphosate, diuron and 3,4-dichloroaniline, showed that it consisted of Arthrobacter sp., Pseudomonas putida, Delftia acidovorans and Brevundimonas sp. strains.
The investigated stormwater basin contains bacterial populations specifically adapted to the transformation of diuron, 3,4-dichloroaniline (3,4-DCA) and glyphosate, and to copper complexation. The mixed culture 106 complexed high amounts of copper ions and degraded glyphosate and diuron without accumulation of the major diuron metabolite 3,4-DCA. Our results also suggest that plants may help to stabilise bacterial-driven pesticide mitigation in environments subject to variable conditions such as stormwater basins.
KeywordsBioremediation Copper Diuron Glyphosate Ribosomal 16S rRNA gene Siderophores
Financial support from the European Union through the project LIFE ENVIRONMENT ArtWET (LIFE 06 ENV/F/000133) “Mitigation of agricultural nonpoint-source pesticides pollution and phytoremediation in artificial wetland ecosystems” (Région Alsace and Conseil Général du Haut-Rhin) is gratefully acknowledged.
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